Various species of Saccharomyces are among the most important industrially grown microorganisms. Long used to leaven bread, produce beer and wine, and as source of food flavorings and micronutrients, these organisms now play a central role in the production of fuel, facilitating the conversion of sugar stocks to ethanol. A metabolically complex organism, yeast is able to grow both aerobically and at least for several generations anaerobically as well. When grown commercially, as in the production of yeast used to support the commercial baking industry, yeasts such as Saccharomyces cerevisiae may be grown in aerated fermentation tanks. The growth of yeast under these conditions may be controlled to increase the production of yeast biomass. One way in which this may be accomplished is to schedule the addition of sugars, such as D-glucose, and the rate of oxygen transfer to the yeast to encourage it to grow aerobically. Various strains of Saccharomyces may also be grown under conditions designed to maximize the production of ethanol. Often times, when the object is to maximize the conversion of sugar to ethanol the level of oxygen in the fermentation vessel may be reduced relative to the levels of oxygen used in the vessel when the object is to maximize yeast biomass production in order to favor anaerobic growth.
Most strains of Saccharomyces have a preference for growth on D-glucose although many strains are known to grow on other naturally occurring hexoses and even some disaccharides as well. The ability of different species of Saccharomyces to grow on different sugars and in the presence of different levels of oxygen accounts for much of its commercial utility including the central role that yeast currently plays in the conversion of plant bio-mass into ethanol for the fuel industry.
One of the best known pathways for the production of ethanol by yeast is the fermentation of 6-carbon sugars (hexoses) into ethanol, especially D-glucose (FIG. 1). One widely used feed stock for the production of ethanol is the polysaccharide starch. Starch is a simple polymer that includes D-glucose. Currently, in the United States at least starch derived from corn is the preferred feed stock for ethanol production by Saccharomyces cerevisiae. Corn is a nutrient-intense crop and currently only the kernels of the corn are a suitable source of starch/D-glucose for ethanol fermentation using yeast. Another source of sugar for the yeast based production of ethanol is sugar cane. Sugar cane is naturally higher in fermentable sugar and may be preferred substrate for the production of ethanol using yeast. However, corn is more widely grown in the United States than is sugar cane. And because of climate it is very likely to remain that way. In any event, the sustainability of corn-based ethanol production has been called into question, and as sugar cane is not a viable option in the United States the bio-fuels industry is looking for other sources of fermentable feed stocks beside corn and sugar cane.
One highly touted feed stock is cellulose, it is considered more sustainable than corn and more readily available than sugar cane. Cellulose processed to produce fermentable sugars may well be the carbon source of choice for the future of ethanol production. Growing yeast in order to increase yeast biomass or to produce ethanol from stocks such as starch or cellulose, requires pre-fermentation processing steps to degrade the bio-polymer cellulose into sugar units, such as D-glucose, maltose, trisaccharides, and tetrasaccharides that can be readily fermented by yeast.
Regardless of its source six-carbon sugars especially D-glucose are the primary energy source for yeast based fermentation. Most species of Saccharomyces that have been characterized grow preferentially on D-glucose. Many of these strains, including many laboratory derived strains of Saccharomyces may grow on hexose sugars other than D-glucose, as well as disaccharides and trisaccharides. However, Saccharomyces preference for growth on D-glucose is so strong that most variants of this yeast including almost all industrially important strains exhibit catabolite repression, that is, the strains will not ferment sugars other than D-glucose so long as there are detectable levels of D-glucose in the feed stock.
The inability of all examined versions of Saccharomyces to vigorously grow on and produce ethanol from sugars other than D-glucose in the presence of D-glucose is unfortunate for the production of yeast biomass and/or ethanol from any feedstock that includes mixtures of fermentable sugars which include D-glucose. For example, D-glucose is liberated by the breakdown of cellulosic biomass into its fermentable components and the presence of D-glucose in the mix of fermentable sugars drastically slows the conversion of the other sugars into ethanol (FIG. 1).
Despite the current technological hurdles to producing ethanol from cellulose the 2007 Energy Independence and Security Act (EISA 2007) mandates that the U.S. rapidly develop technologies to produce cellulosic ethanol to displace imported petroleum. Accordingly, there is a need for novel strains of industrial Saccharomyces and for methods of creating these industrial strains that readily convert sugars other than just D-glucose into biomass or ethanol even in the presence of significant amounts of D-glucose. Some aspects of the present invention address these needs.